Carl S. Thummel

The nuclear receptor superfamily: The second decade

David J. Mangelsdorf,’ Carl Thummel,2 Miguel Beato,3 Peter Herrlich,4 Giinther Schiitq5 Kazuhiko Umesono,6 Bruce Blumberg,’ Philippe Kastner,’ Manuel Mark,* Pierre Chambon,8 and Ronald M. Evan&‘* ‘Howard Hughes Medical Institute University of Texas Southwestern Medical Center Dallas, Texas 75235-9050 *Howard Hughes Medical Institute University of Utah Salt Lake City, Utah 84112 31nstutut fiir Molekularbiologie und Tumorforschung 35037 Marburg Federal Republic of Germany 4Forschungszentrum Karlsruhe lnstitut Genetik 76021 Karlsruhe Federal Republic of Germany 5Deutsches Krebsforschungszentrum 69120 Heidelberg Federal Republic of Germany GAdvanced Institute of Science and Technology Graduate School of Biological Sciences Nara 630-01 Japan ‘The Salk Institute for Biological Studies La Jolla, California 92037-5800 Blnstitut de Genetique et de Biologie Moleculaire et Cellulaire Centre National de la Recherche Scientifique lnstitut National de la Sante et de la Recherche M6dicale 67404 lllkirch Cedex Strasbourg France gHoward Hughes Medical Institute The Salk Institute for Biological Studies La Jolla, California 92037-5800

Vectors for Drosophila P-element-mediated transformation and tissue culture transfection

We describe nine P-element vectors that can be used to study gene regulation and function in Drosophila. These vectors were designed for use in germline transformation and cell culture transfection assays. One set consists of five P elements that can be used to study transcriptional regulatory sequences. These vectors contain several unique restriction sites for insertion of a foreign promoter upstream from either a cat or lacZ reporter gene. Two of the beta-galactosidase-coding vectors also require the insertion of a start codon for translation of the reporter enzyme and thus can be used to study translational regulatory sequences. The second set of P elements consists of four vectors that contain the Drosophila cytoplasmic actin 5C promoter and polyadenylation signals. Upon insertion of a foreign DNA segment, these vectors direct constitutive expression of the encoded RNA and protein.

Flies on steroids — Drosophila metamorphosis and the mechanisms of steroid hormone action

Recent studies have provided new insights into the molecular mechanisms by which the steroid hormone ecdysone triggers the larval-to-adult metamorphosis of Drosophila. Ecdysone-induced transcription factors activate large sets of secondary-response genes and provide the competence for subsequent regulatory responses to the hormone. It seems likely that similar hormone-triggered regulatory hierarchies exist in other higher organisms and that Drosophila is providing our first glimpses of the complexities of these gene networks.

NUCLEAR RECEPTORS — A PERSPECTIVE FROM DROSOPHILA

Nuclear receptors are ancient ligand-regulated transcription factors that control key metabolic and developmental pathways. The fruitfly Drosophila melanogaster has only 18 nuclear-receptor genes — far fewer than any other genetic model organism and representing all 6 subfamilies of vertebrate receptors. These unique attributes establish the fly as an ideal system for studying the regulation and function of nuclear receptors during development. Here, we review recent breakthroughs in our understanding of D. melanogaster nuclear receptors, and interpret these results in light of findings from their evolutionarily conserved vertebrate homologues.

From embryogenesis to metamorphosis: the regulation and function of Drosophila nuclear receptor superfamily members.

The discovery of the nuclear receptor superfamily and detailed studies of receptor function have revolutionized our understanding of hormone action. Studies of nuclear receptor superfamily members in the fruit fly, Drosophila melanogaster, have contributed to these breakthroughs by providing an ideal model system for defining receptor function in the context of a developing animal. To date, 16 genes of the nuclear receptor superfamily have been isolated in Drosophila, all encoding members of the heterodimeric and orphan classes of receptors (reviewed by Mangelsdorf and Evans, 1995 [this issue of Ce//]). Some superfamily members appear to function specifically during embryonic stages while others are required during multiple stages of development. Remarkably, half of the known Drosophila superfamily members are regulated by the steroid hormone ecdysone and appear to contribute to the complex developmental pathways associated with metamorphosis. This review provides a brief survey of our current understanding of how nuclear receptor superfamily members function during Drosophila development and how their activities relate to those of their vertebrate homologs.

Nuclear receptors |[mdash]| a perspective from Drosophila

Nuclear receptors are ancient ligand-regulated transcription factors that control key metabolic and developmental pathways. The fruitfly Drosophila melanogaster has only 18 nuclear-receptor genes — far fewer than any other genetic model organism and representing all 6 subfamilies of vertebrate receptors. These unique attributes establish the fly as an ideal system for studying the regulation and function of nuclear receptors during development. Here, we review recent breakthroughs in our understanding of D. melanogaster nuclear receptors, and interpret these results in light of findings from their evolutionarily conserved vertebrate homologues.

A Mitochondrial Pyruvate Carrier Required for Pyruvate Uptake in Yeast, Drosophila, and Humans

Letting Pyruvate In Transport of pyruvate is an important event in metabolism whereby the pyruvate formed in glycolysis is transported into mitochondria to feed into the tricarboxylic acid cycle (see the Perspective by Murphy and Divakaruni). Two groups have now identified proteins that are components of the mitochondrial pyruvate transporter. Bricker et al. (p. 96, published online 24 May) found that the proteins mitochondrial pyruvate carrier 1 and 2 (MPC1 and MPC2) are required for full pyruvate transport in yeast and Drosophila cells and that humans with mutations in MPC1 have metabolic defects consistent with loss of the transporter. Herzig et al. (p. 93, published online 24 May) identified the same proteins as components of the carrier in yeast. Furthermore, expression of the mouse proteins in bacteria conferred increased transport of pyruvate into bacterial cells. The genes encoding two components of the pyruvate transporter in mitochondria have been identified. Pyruvate constitutes a critical branch point in cellular carbon metabolism. We have identified two proteins, Mpc1 and Mpc2, as essential for mitochondrial pyruvate transport in yeast, Drosophila, and humans. Mpc1 and Mpc2 associate to form an ~150-kilodalton complex in the inner mitochondrial membrane. Yeast and Drosophila mutants lacking MPC1 display impaired pyruvate metabolism, with an accumulation of upstream metabolites and a depletion of tricarboxylic acid cycle intermediates. Loss of yeast Mpc1 results in defective mitochondrial pyruvate uptake, and silencing of MPC1 or MPC2 in mammalian cells impairs pyruvate oxidation. A point mutation in MPC1 provides resistance to a known inhibitor of the mitochondrial pyruvate carrier. Human genetic studies of three families with children suffering from lactic acidosis and hyperpyruvatemia revealed a causal locus that mapped to MPC1, changing single amino acids that are conserved throughout eukaryotes. These data demonstrate that Mpc1 and Mpc2 form an essential part of the mitochondrial pyruvate carrier.

A Steroid-Triggered Transcriptional Hierarchy Controls Salivary Gland Cell Death during Drosophila Metamorphosis

The steroid hormone ecdysone signals the stage-specific programmed cell death of the larval salivary glands during Drosophila metamorphosis. This response is preceded by an ecdysone-triggered switch in gene expression in which the diap2 death inhibitor is repressed and the reaper (rpr) and head involution defective (hid) death activators are induced. Here we show that rpr is induced directly by the ecdysone-receptor complex through an essential response element in the rpr promoter. The Broad-Complex (BR-C) is required for both rpr and hid transcription, while E74A is required for maximal levels of hid induction. diap2 induction is dependent on betaFTZ-F1, while E75A and E75B are each sufficient to repress diap2. This study identifies transcriptional regulators of programmed cell death in Drosophila and provides a direct link between a steroid signal and a programmed cell death response.

Temporal coordination of regulatory gene expression by the steroid hormone ecdysone.

In Drosophila, pulses of the steroid hormone ecdysone function as temporal signals that trigger the major postembryonic developmental transitions. The best characterized of these pulses activates a series of puffs in the polytene chromosomes as it triggers metamorphosis. A small set of early puffs is induced as a primary response to the hormone. These puffs encode regulatory proteins that both repress their own expression and activate a large set of late secondary response genes. We have used Northern blot analysis of RNA isolated from staged animals and cultured organs to study the transcription of three primary response regulatory genes, E75, BR‐C and EcR. Remarkably, their patterns of transcription in late larvae can be defined in terms of two responses to different ecdysone concentrations. The class I transcripts (E74B and EcR) are induced in mid‐third instar larvae in response to the low, but increasing, titer of ecdysone. As the hormone concentration peaks in late third instar larvae, these transcripts are repressed and the class II RNAs (E74A, E75A and E75B) are induced. The BR‐C RNAs appear to have both class I and class II characteristics. These data demonstrate that the relatively simple profile of a hormone pulse contains critical temporal information that is transduced into waves of primary response regulatory gene activity.

Molecular mechanisms of developmental timing in C. elegans and Drosophila.

Characterization of the heterochronic genes has provided a strong foundation for understanding the molecular mechanisms of developmental timing in C. elegans. In apparent contrast, studies of developmental timing in Drosophila have demonstrated a central role for gene cascades triggered by the steroid hormone ecdysone. In this review, I survey the molecular mechanisms of developmental timing in C. elegans and Drosophila and outline how common regulatory pathways are beginning to emerge.